See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/327106025 Resistance of cyanobacteria to space and Mars environments, in the frame of the EXPOSE-R2 space mission and beyond Thesis · April 2018 DOI: 10.13140/RG.2.2.28437.88808 CITATIONS READS 3 522 1 author: Cyprien Verseux ZARM 36 PUBLICATIONS 413 CITATIONS SEE PROFILE Some of the authors of this publication are also working on these related projects: HI-SEAS IV Crew View project BIOMEX (Biology and Mars Experiment) View project All content following this page was uploaded by Cyprien Verseux on 11 September 2019. The user has requested enhancement of the downloaded file. UNIVERSITY OF ROME "TOR VERGATA" DOCTORAL PROGRAM IN CELLULAR AND MOLECULAR BIOLOGY XXIXth CYCLE FACULTY OF MATHEMATICAL, PHYSICAL AND NATURAL SCIENCES DEPARTMENT OF BIOLOGY Resistance of cyanobacteria to space and Mars environments, in the frame of the EXPOSE-R2 space mission and beyond Cyprien Verseux Y.Y. 2016/2017 (defended on 04/27/2018) Co-Supervisor: Pr. Daniela Billi Co-Supervisor: Dr. Lynn Rothschild Tutor: Pr. Manuela Helmer Citterich Program Coordinator: Pr. Francesco Cecconi 1 2 TABLE OF CONTENTS Summary of doctoral research activities ........................................................ 7 PART I: RESISTANCE OF CHROOCOCCCIDIOPSIS SPP. TO SPACE AND MARS-LIKE CONDITIONS ............................................................. 11 Chapter 1: Introducing Part I ........................................................................ 13 Cyanobacteria and the search for life beyond Earth ................................. 13 Exposure of cyanobacteria to space and Mars-like conditions ................. 26 Aim of Part I and chapters’ description .................................................... 34 Chapter 2: Survival and biosignature detection of Chroococcidiopsis spp. after ground-based simulations of extraterrestrial environments .......................... 41 Introduction .............................................................................................. 42 Material and Methods ............................................................................... 45 Cyanobacterial strains and culture conditions ...................................... 45 Sample preparation ............................................................................... 45 Test facilities and exposure conditions ................................................. 48 Survival ................................................................................................ 49 Random amplification of polymorphic DNA (RAPD) assay ............... 49 Confocal laser scanning microscopy .................................................... 50 Membrane damage assessment ............................................................. 50 Results ...................................................................................................... 52 Survival ................................................................................................ 52 DNA detection by PCR-based assay .................................................... 53 Emission spectra of photosynthetic pigments....................................... 55 Membrane damage assessment ............................................................. 61 Discussion ................................................................................................ 62 Exposure conditions ............................................................................. 62 3 Survival and DNA preservation ........................................................... 64 Preservation of pigments’ autofluorescence ......................................... 66 Potential mechanisms behind Chroococcidiopsis’s spp. resistance ..... 67 Conclusion ............................................................................................ 70 Chapter 3: Enhanced resistance of Chroococcidiopsis biofilms, compared to their planktonic counterparts, to space and simulated Martian conditions in low Earth orbit .............................................................................................. 73 Introduction .............................................................................................. 74 Materials and methods .............................................................................. 77 Organisms and sample preparation....................................................... 77 Exposure in low Earth orbit .................................................................. 77 Mission ground references and laboratory controls .............................. 79 Survival ................................................................................................ 80 qPCR assays ......................................................................................... 80 Confocal laser scanning microscopy .................................................... 80 Results ...................................................................................................... 83 Survival ................................................................................................ 83 DNA damage revealed by qPCR .......................................................... 84 Confocal laser scanning microscopy .................................................... 87 Discussion ................................................................................................ 91 Differences in survival ......................................................................... 91 Differences in DNA damage ................................................................ 94 Preservation of photosynthetic pigments .............................................. 97 Acknowledgements .................................................................................. 99 Chapter 4: Responses of the desert cyanobacterium Chroococcidiopsis to simulated Martian conditions in low Earth orbit: implications for the limits of terrestrial life and the habitability of Mars ................................................. 101 Introduction ............................................................................................ 102 Materials and methods ............................................................................ 105 4 Mars regolith analogues ..................................................................... 105 Organisms and sample preparation..................................................... 105 Exposure in low Earth orbit ................................................................ 106 Ground references and laboratory controls ........................................ 108 Colony forming ability assays ............................................................ 109 Random Amplification of Polymorphic DNA .................................... 109 Confocal laser scanning microscopy .................................................. 109 Results ................................................................................................ 17411 Survival .............................................................................................. 111 DNA damage as revealed by random amplification of polymorphic DNA ............................................................................................................ 111 Alteration of photosynthetic pigments as revealed by CLSM ............ 113 Discussion .............................................................................................. 118 Survival .............................................................................................. 118 DNA damage as revealed by Random Amplification of Polymorphic DNA ................................................................................................... 121 Alteration of photosynthetic pigments as revealed by CLSM ............ 121 Conclusions ........................................................................................ 124 Acknowledgements ................................................................................ 125 PART II: CYANOBACTERIUM-BASED LIFE-SUPPORT SYSTEMS FOR THE FUTURE OF CREWED SPACE EXPLORATION.......................... 127 Chapter 5: Introducing Part II ..................................................................... 129 Aim of Part II and chapter’s description .............................................. 1677 Chapter 6: Cyanobacterium biomass as a substrate for heterotrophic growth on Mars ....................................................................................................... 169 Introduction ............................................................................................ 169 Materials and methods ............................................................................ 171 Bacterial cultures ................................................................................ 171 5 Lithotrophic growth of Anabaena sp. PCC 7120 with Mars regolith simulant .............................................................................................. 171 Cyanobacterium biomass as a nutrient source for heterotrophic bacteria ............................................................................................................ 171 Characterization of the cyanobacterium medium ............................... 172 Perchlorate growth assays .................................................................. 172 Results .................................................................................................... 174 Lithotrohic